An essential step toward understanding the relationship between genotype and phenotype will be to determine how frequently genes interact with each other and with the environments in which they are expressed. Both types of interactions are known to occur for specific genes and conditions; however, it is unclear how generally genetic and environmental background influence gene expression and function. In the proposed project, Escherichia coli and the genetic tools that have been developed for this model organism will be used to investigate the effects of these interactions on fitness. Random transposon insertion mutants and their pairwise double mutant genotypes will be constructed using transposon mutagenesis and P1 transduction. The fitness of all genotypes will be evaluated, and the presence and nature of fitness effects due to 'gene-gene' interactions (epistatis) will be determined. Fitnesses will be quantified in a range of environments, so that 'gene- environment' and 'gene-gene-environment' interactions can be measured. The mutated genes will be identified by sequencing bacterial DNA adjacent to the transposons using transposon-specific primers. Genes will be assigned to functional classes by comparison with the published complete sequence. The relationship between the functional classes of affected genes and the types of interactions in which they were detected will then be analyzed. Exploring genes' interactions with each other and with the environments in which they are expressed will increase our understanding of how classes of genes behave.